Detection method and application of Guanxinjing tablet

Abstract

The invention relates to the technical field of pharmacy, in particular to a detection method and application of Guanxinjing tablets. The detection method comprises the following steps: (1) identifying paeoniflorin in the Guanxinjing tablet by a thin layer chromatography, and (2) detecting the contents of tanshinol, protocatechualdehyde and salvianolic acid B in the Guanxinjing tablet by a high performance liquid chromatography, the method for detecting the content of each drug component in the monarch drug salvia miltiorrhiza in the Guanxinjing tablet and the method for detecting the drug components in the ministerial drug in the Guanxinjing tablet are not disclosed in the prior art, and a basis is provided for more accurate detection of the Guanxinjing tablet. The method for detecting paeoniflorin in radix paeoniae rubra through thin layer chromatography is good in specificity and durability. The high performance liquid chromatography for detecting the contents of tanshinol, protocatechuic aldehyde and salvianolic acid B in salvia miltiorrhiza is good in specificity, the recovery rate ranges from 95% to 105%, and the high performance liquid chromatography is good in accuracy, precision, linear relation and durability. And a basis is provided for strictly controlling the quality of the coronary heart calming tablet.

Description

Technical Field

[0001] The present invention relates to the technical field of pharmaceuticals, and particularly relates to a detection method and application of Guanxinjing tablets. Background Art

[0002] Guanxinjing tablets are based on Xuefu Zhuyu Decoction in "Corrections to Medical Writings" by Wang Qingren, a medical scientist in the Qing Dynasty, and are developed by adding and subtracting various herbs in the formula according to traditional Chinese medicine theory. The final formula of Guanxinjing tablets is: the monarch herb is Salvia miltiorrhiza; the minister herbs are Paeonia lactiflora, Ligusticum chuanxiong, Carthamus tinctorius, and Panax ginseng; the assistant herbs are Polygonatum odoratum and Notoginseng; the guiding herbs are Styrax and Borneol. The quality of Guanxinjing tablets prepared by different preparation methods is different. Therefore, it is very important to identify the quality of Guanxinjing tablets.

[0003] In the existing standard with the standard number WS3 - B - 3439 - 98, in addition to the detection of the conventional items of tablets, there is only the thin - layer identification of Ligusticum chuanxiong and panaxadiol, without the identification of other components and even less the determination of content. In order to strictly control the quality of Guanxinjing tablets, it is necessary to accurately identify other components therein.

[0004] Based on this, the present invention is proposed. Summary of the Invention

[0005] The purpose of the present invention is to provide a method for accurately identifying the quality of the monarch herb and minister herbs in Guanxinjing tablets, and particularly relates to a detection method and application of Guanxinjing tablets.

[0006] In order to achieve the above - mentioned invention purpose, the present invention provides the following technical solutions:

[0007] The present invention provides a detection method for Guanxinjing tablets, wherein the Guanxinjing tablets include the monarch herb Salvia miltiorrhiza and the minister herb Paeonia lactiflora, and the detection method includes the following steps:

[0008] (1) Identifying paeoniflorin in Guanxinjing tablets by thin - layer chromatography

[0009] 1.1 Extract the active ingredients from Guanxinjing tablets and Paeonia lactiflora respectively by the same method, prepare them into solutions to obtain the test solution and the solution of the control medicinal material;

[0010] 1.2 Mix paeoniflorin reference substance with methanol to obtain the reference substance solution;

[0011] 1.3 Take the test solution, the solution of the control medicinal material, and the reference substance solution and spot them on the same silica gel G thin - layer plate respectively, develop with the developing agent, take out, dry, spray the color - developing agent, and heat for color development; judge whether the test solution contains paeoniflorin according to the color - developing position and color of the solution of the control medicinal material and the reference substance solution;

[0012] Paeoniflorin in Guanxinjing tablets belongs to Paeonia lactiflora;

[0013] (2) Determination of the Contents of Danshensu, Protocatechuic Aldehyde and Salvianolic Acid B in Guanxinjing Tablets by High Performance Liquid Chromatography

[0014] 2.1 Take the reference substances of danshensu sodium, protocatechuic aldehyde and salvianolic acid B, mix them with 65 - 75 vt% methanol to obtain a reference substance solution.

[0015] 2.2 Extract the active ingredients in Guanxinjing Tablets, prepare them into a solution to obtain a test sample solution.

[0016] 2.3 Inject the reference substance solution and the test sample solution into a liquid chromatograph for determination respectively.

[0017] The chromatographic conditions are as follows: the chromatographic column is a C18 column (4.6 mm × 250 mm, 5 μm); acetonitrile is used as mobile phase A, and 0.05 - 0.15% trifluoroacetic acid is used as mobile phase B, the column temperature is 28 - 32 °C, the detection wavelength is 260 - 290 nm, and the number of theoretical plates calculated based on the salvianolic acid B peak is not less than 30000.

[0018] The elution conditions are as follows: 0 - 20 min, 5% mobile phase A, 95% mobile phase B; 20 - 30 min, 5 → 9% mobile phase A, 95 → 91% mobile phase B; 30 - 35 min, 9% mobile phase A, 91% mobile phase B; 35.1 - 95 min, 20% mobile phase A, 80% mobile phase B.

[0019] Danshensu, protocatechuic aldehyde and salvianolic acid B in Guanxinjing Tablets belong to Salvia miltiorrhiza.

[0020] Preferably, the method for extracting the active ingredients in Guanxinjing Tablets and Paeonia lactiflora Pall. in step 1.1 is as follows:

[0021] Crush Guanxinjing Tablets or Paeonia lactiflora Pall. to obtain a powder; mix the powder with methanol and ultrasonicate for 25 - 35 min, filter, and take the filtrate; concentrate the filtrate to obtain a concentrated solution; mix the concentrated solution with neutral alumina, evaporate to dryness to obtain a dried product; add the dried product into a neutral alumina column, elute, collect the eluate to obtain the active ingredients.

[0022] Evaporate the active ingredients to dryness to obtain a residue, dissolve the residue in methanol, and prepare a test sample solution and a control crude drug solution respectively.

[0023] The mass - volume ratio of the powder to methanol is 1 - 1.5 g:25 mL.

[0024] The mass - volume ratio of neutral alumina to the concentrated solution is 0.8 - 1.2 g:0.8 - 1.2 mL.

[0025] The particle size of the neutral alumina is 200 - 300 mesh.

[0026] The particle size of the neutral alumina contained in the neutral alumina column is 200 - 300 mesh, and the inner diameter of the neutral alumina column is 0.8 - 1.2 cm;

[0027] The solution during elution is methanol.

[0028] Preferably, the mass - volume ratio of the paeoniflorin reference substance to methanol in step 1.2 is 0.8 - 1.2 mg: 0.8 - 1.2 mL.

[0029] Preferably, the developing agent in step 1.3 comprises components in the following volume ratio:

[0030] Chloroform: methanol: water is 38 - 42: 8 - 12: 1;

[0031] The color - developing agent in step 1.3 is sulfuric acid - ethanol solution; the concentration of sulfuric acid contained in the sulfuric acid - ethanol solution is 10 vt%.

[0032] The temperature for heating and color - developing in step 1.3 is 100 - 110 °C.

[0033] Preferably, the concentration of sodium danshensu in the reference substance solution of step 2.1 is 0.1 - 0.14 mg / mL, the concentration of protocatechuic aldehyde is 10 - 14 μg / mL, and the concentration of salvianolic acid B is 0.26 - 0.30 mg / mL.

[0034] Preferably, the method for extracting the active ingredients from Guanxinjing tablets in step 2.2 is as follows:

[0035] Crush Guanxinjing tablets to obtain powder; mix the powder with methanol to obtain a mixture, and weigh the mixture; heat the mixture under reflux for 25 - 35 min, cool to obtain the active ingredients;

[0036] Weigh the obtained active ingredients; make up the weight of the active ingredients to the same as that of the mixture with methanol, filter, take the filtrate to obtain the test solution;

[0037] The mass - volume ratio of the powder to methanol is 0.4 - 0.6 g: 25 mL.

[0038] The present invention also provides the application of the described detection method in accurately identifying whether paeoniflorin is contained in Guanxinjing tablets.

[0039] The present invention also provides the application of the described detection method in accurately detecting the content of sodium danshensu in Guanxinjing tablets.

[0040] The present invention also provides the application of the described detection method in accurately detecting the content of protocatechuic aldehyde in Guanxinjing tablets.

[0041] The present invention also provides an application of the detection method in accurately detecting the content of salvianolic acid B in Guanxinjing tablets.

[0042] The present invention provides a detection method for Guanxinjing tablets. The detection method of the present invention includes: (1) identifying paeoniflorin in Guanxinjing tablets by thin layer chromatography; (2) detecting the contents of danshensu, protocatechuic aldehyde and salvianolic acid B in Guanxinjing tablets by high performance liquid chromatography, filling the gaps in the prior art where there is no disclosure on how to detect the contents of each drug component in the monarch drug salvia miltiorrhiza and the drug components in the ministerial drugs in Guanxinjing tablets, and providing a basis for more accurately detecting Guanxinjing tablets.

[0043] The method for detecting paeoniflorin in radix paeoniae rubra by the thin layer chromatography method of the present invention has good specificity and good durability. The method for detecting the contents of danshensu, protocatechuic aldehyde and salvianolic acid B in salvia miltiorrhiza by high performance liquid chromatography has good specificity, with a recovery rate between 95% and 105%, good accuracy, good precision, good linear relationship and good durability, providing a basis for strictly controlling the quality of Guanxinjing tablets. Description of the Drawings

[0044] Figure 1 Chromatogram of Guanxinjing tablets without film coating;

[0045] Figure 2 Chromatogram of Guanxinjing tablets with film coating removed;

[0046] Figure 3 Chromatogram of the negative sample of salvia miltiorrhiza in Guanxinjing tablets;

[0047] Figure 4 Chromatogram of Guanxinjing tablets;

[0048] Figure 5 Standard curve of sodium danshensu;

[0049] Figure 6 Standard curve of protocatechuic aldehyde;

[0050] Figure 7 Standard curve of salvianolic acid B. Detailed Embodiments

[0051] The solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0052] Unless otherwise specified, the Guanxinjing tablets described in the embodiments of the present invention are all film-coated Guanxinjing tablets produced by Baoding Traditional Chinese Medicine Pharmaceutical Co., Ltd., with an average tablet weight of 0.31 g / tablet.

[0053] Preparation method of film-coated Guanxinjing tablets:

[0054] Use a film coating premix (transparent white) to prepare the isolating layer coating solution, and use a film coating premix (brownish red) to prepare the color layer coating solution. Start the high-efficiency coating machine, first coat the core tablets of Guanxinjing Tablets with the isolating layer, and then coat the color layer. The weight gain of the core tablets after film coating does not exceed 4%.

[0055] The transparent white film coating premix and the brownish red film coating premix are purchased from Beijing Yingmao Pharmaceutical Co., Ltd.

[0056] The paeoniflorin, danshensu sodium, protocatechuic aldehyde, and salvianolic acid B reference substances in the examples of the present invention are all purchased from the National Institutes for Food and Drug Control.

[0057] The high-performance liquid chromatograph in the examples of the present invention is Shimadzu LC-20AD XR , and the detector is Shimadzu SPD-20A ultraviolet detector.

[0058] In the present invention, the molecular weight of danshensu sodium (C9H9O5Na) is 220.16, and the molecular weight of danshensu (C9H 10 O5) is 198.17; using danshensu sodium as a reference to detect the content of danshensu in Guanxinjing Tablets is because danshensu sodium is more stable (danshensu is easily oxidized) and is soluble in water. After detecting the content of danshensu sodium, according to the different molecular weights of danshensu and danshensu sodium, the content of danshensu in Guanxinjing Tablets can be calculated.

[0059] The content of danshensu = (the content of danshensu sodium × the molecular weight of danshensu / the molecular weight of danshensu sodium) × 100%.

[0060] Example 1

[0061] Identification of paeoniflorin in Guanxinjing Tablets by thin layer chromatography

[0062] 1.1 Detection method

[0063] Take 10 tablets of Guanxinjing Tablets with the product batch number 250201, grind them finely, weigh 1.5 g, add 25 mL of methanol, ultrasonically treat for 30 min, filter, concentrate the filtrate to 1 mL to obtain a concentrated solution. Add 1 g of neutral alumina (200 - 300 mesh) to the concentrated solution, mix well, evaporate to dryness to obtain a dried product. Add the dried product to a neutral alumina column (200 - 300 mesh, 1 g, inner diameter 1 cm), elute with 20 mL of methanol, collect the eluate, evaporate to dryness, and dissolve the residue in 1 mL of methanol to obtain a test solution.

[0064] Take 1 g of the reference crude drug of Paeonia lactiflora Pall., and prepare a reference crude drug solution in the same way.

[0065] Take paeoniflorin reference substance, add methanol to make a solution containing 1 mg per 1 mL as the reference substance solution. According to the thin-layer chromatography method (General Principles 0502 of Chinese Pharmacopoeia 2020 Edition), draw 3 μL of each of the above three solutions, spot them on the same silica gel G thin-layer plate respectively, use the developing agent prepared by mixing chloroform: methanol: water in a volume ratio of 40:10:1 for development, take out, dry in air, spray with 10% sulfuric acid ethanol solution, and heat at 105 °C until the spots are clearly visible. As a result, in the chromatogram of the test sample, at the positions corresponding to the chromatograms of the reference medicinal material and the reference substance, spots of the same color appear. It shows that paeoniflorin is contained in Guanxinjing tablets.

[0066] 1.2 Methodology investigation

[0067] 1.2.1 Specificity

[0068] Take 20 Guanxinjing tablets with the product batch number 250201, and test 10 of them according to the test method in 1.1. For the other 10 tablets, after removing the film coating, test them according to the test method in 1.1;

[0069] Prepare Guanxinjing tablets without Paeonia lactiflora according to the prescription process for preparing Guanxinjing tablets as the negative sample, and test them according to the test method in 1.1.

[0070] The results show that the thin-layer chromatography spots of Guanxinjing tablets with and without film coating are clear, and there is no difference in the thin-layer chromatography spots. Whether to remove the film coating has no influence on the identification. At the same time, no chromatographic spots appear at the positions corresponding to the reference substance chromatogram in the negative sample, indicating that other drug components and excipients in Guanxinjing tablets have no interference in the detection of paeoniflorin in Paeonia lactiflora, indicating that the detection method has good specificity. By detecting paeoniflorin, it can be determined whether Paeonia lactiflora is contained in Guanxinjing tablets.

[0071] 1.2.2 Durability

[0072] Use silica gel G thin-layer plates from different manufacturers and test according to the test method in 1.1. The manufacturers of silica gel G thin-layer plates include Qingdao Ocean Chemical Co., Ltd., Yantai Jiangyou Silica Gel Development Co., Ltd., and Hunan Bikeman Holdings Co., Ltd.

[0073] The test results show that when using silica gel G thin-layer plates from different manufacturers for detection, the spots are clear, indicating that the detection method of the present invention has good durability.

[0074] Example 2

[0075] Determination of the contents of danshensu, protocatechuic aldehyde and salvianolic acid B in Guanxinjing tablets by high performance liquid chromatography

[0076] 2.1 Detection method

[0077] Preparation of reference solution: Weigh an appropriate amount of sodium danshensu, protocatechuic aldehyde, and salvianolic acid B reference substances accurately, dissolve them in 70% methanol to prepare a mixed solution containing 0.12 mg of sodium danshensu, 12 μg of protocatechuic aldehyde, and 0.28 mg of salvianolic acid B per 1 mL. That is how you get it.

[0078] Preparation of test solution: Take 10 Guanjingxin Tablets with the product batch number 250201, grind them into fine powder, take 0.5 g, weigh it accurately, place it in a stoppered conical flask, accurately add 25 mL of 70% methanol, weigh it, heat under reflux for 30 min, cool, weigh again, make up the lost weight with 70% methanol, shake well, filter, and take the filtrate. That is how you get it.

[0079] Determination method: Accurately pipette 10 μL of the reference solution and the test solution respectively, inject them into the liquid chromatograph for determination. That is how you get the results.

[0080] Chromatographic conditions: The chromatographic column is a Waters Symmetry C18 column (4.6 mm × 250 mm, 5 μm); acetonitrile is used as mobile phase A, and 0.1% trifluoroacetic acid is used as mobile phase B. The column temperature is 30 °C, the detection wavelength is 281 nm, and the number of theoretical plates calculated based on the salvianolic acid B peak is not less than 30000. The elution conditions are shown in Table 1.

[0081] Table 1 Elution conditions

[0082] Time (min) Mobile Phase A (%) Mobile Phase B (%) 0～20 5 95 20～30 5→9 95→91 30～35 9 91 35.1～95 20 80

[0083] Requirement: During detection, the total amount of sodium danshensu, protocatechuic aldehyde, and salvianolic acid B in each Guanjingxin Tablet shall not be less than 1.0 mg, and the content of salvianolic acid B shall not be less than 0.7 mg.

[0084] 2.2 Methodological investigation

[0085] 2.2.1 Specificity

[0086] Take 20 Guanjingxin Tablets with the product batch number 250201, 10 of which do not remove the film coating, and the other 10 remove the film coating. Conduct the test according to the test method in 2.1 respectively. The results are as Figures 1 - 2 shown.

[0087] The results show that there is no difference in the chromatographic peaks whether the film coating is removed or not. The retention times of the target chromatographic peaks of the test sample and the reference sample are the same, and the number of theoretical plates of salvianolic acid B is greater than 30000. The specificity of this method is good. The three chromatographic peaks marked with retention times in the figure are sodium danshensu, protocatechuic aldehyde, and salvianolic acid B in sequence.

[0088] Prepare Guanjingxin Tablets without danshen according to the prescription process of Guanjingxin Tablets as a negative sample, compare it with Guanjingxin Tablets, and conduct the test according to the test method in 2.1. The results are as Figures 3 - 4as shown

[0089] As a result, there were no chromatographic peaks at the same position in the chromatogram of the negative sample of Guanxinjing Tablets, proving that other medicinal flavors in the prescription of Guanxinjing Tablets did not interfere with the determination of danshensu, protocatechuic aldehyde, and salvianolic acid B. Figure 4 The chromatographic peaks in it were successively indicated as sodium danshensu, protocatechuic aldehyde, and salvianolic acid B.

[0090] 2.2.2 Accuracy

[0091] Prepare the reference substance solution according to the test method in 2.1.

[0092] Take Guanxinjing Tablets with known danshensu (0.7819 mg / tablet), protocatechuic aldehyde (0.07443 mg / tablet), and salvianolic acid B (1.9958 mg / tablet), and prepare the test solution according to the detection method in 2.1.

[0093] Take 5 mL of the reference substance solution and 5 mL of the test solution and mix them as the mixed solution for accuracy testing.

[0094] Precisely weigh 10 μL of the reference substance solution and 10 μL of the mixed solution respectively, inject them into the liquid chromatograph, and conduct detection according to the detection method in 2.1, with 6 parallel tests. Calculate the recovery rate. The results are shown in Tables 2 - 4.

[0095] Table 2 Recovery rate of danshensu

[0096]

[0097]

[0098] Table 3 is the recovery rate of protocatechuic aldehyde

[0099]

[0100] Table 4 is the recovery rate of salvianolic acid B

[0101]

[0102] Recovery rate = (C - A) / B × 100%.

[0103] As a result, the average recovery rate of danshensu in the test solution was 101.05%, and the RSD of the recovery rate was 0.3%; the average recovery rate of protocatechuic aldehyde was 97.29%, and the RSD of the recovery rate was 0.2%; the average recovery rate of salvianolic acid B was 98.73%, and the RSD of the recovery rate was 0.4%; the accuracy of this method was good.

[0104] 2.2.3 Precision

[0105] According to the inspection method in 2.1, prepare the reference solution and 6 portions of test solutions; precisely pipette 10 μl of the reference solution, inject it into the liquid chromatograph, and inject continuously for 5 times; precisely pipette 10 μl of the test solution, inject it into the liquid chromatograph, and determine. The results are shown in Tables 5 - 7.

[0106] Table 5 Table of peak areas of the reference solution

[0107]

[0108] Table 6 shows the contents of danshensu, protocatechuic aldehyde, and salvianolic acid B in the test solution

[0109]

[0110] Table 7 shows the total contents of danshensu, protocatechuic aldehyde, and salvianolic acid B in the test solution

[0111]

[0112]

[0113] The results show that the RSDs of the peak areas of sodium danshensu, protocatechuic aldehyde, and salvianolic acid B in the reference solution are 0.2%, 0.2%, and 0.2% respectively; the average content of danshensu in the 6 portions of test samples is 0.7819 mg / tablet, and the RSD of the content is 0.4%; the average content of protocatechuic aldehyde is 0.0744 mg / tablet, and the RSD of the content is 0.9%; the average content of salvianolic acid B is 1.9958 mg / tablet, and the RSD of the content is 0.8%; the average total content of the three components is 2.8522 mg / tablet, and the RSD of the total content is 0.5%; the precision of this method is good.

[0114] 2.2.4 Linear relationship

[0115] Precisely weigh appropriate amounts of sodium danshensu reference substance, protocatechuic aldehyde reference substance, and salvianolic acid B reference substance, and prepare a mixed solution with 70% methanol (the concentrations of sodium danshensu, protocatechuic aldehyde, and salvianolic acid B in the mixed solution are approximately 600 μg / mL, 60 μg / mL, and 1400 μg / mL), and dilute according to Table 8 to prepare the reference mixed solution.

[0116] Table 8 Sampling of the reference mixed solution

[0117] Initial Concentration Aliquot Volume (mL) 3 3 2 5 3 Diluted Volume (mL) 25 20 10 20 10

[0118] Precisely pipette 10 μl of the reference solutions with different concentrations above, inject them into the liquid chromatograph, and determine according to the detection method in 2.1. Use the injection amount of the reference substance (μg) as the abscissa and the peak area as the ordinate to plot the standard curve, and calculate the regression equation and correlation coefficient. The relevant data and results are shown in Table 9 andFigures 5 - 7 as shown

[0119] Table 9 Linear Equation

[0120]

[0121]

[0122] The results showed that danshensu sodium had a good linear relationship between 72 μg / mL and 180 μg / mL, with a correlation coefficient of 0.9999; protocatechuic aldehyde had a good linear relationship between 7.61 μg / mL and 19.03 μg / mL, with a correlation coefficient of 0.9983; salvianolic acid B had a good linear relationship between 168.19 μg / mL and 420.47 μg / mL, with a correlation coefficient r of 0.9999; the method had good linearity.

[0123] 2.2.5 Range

[0124] According to the test method in 2.1, prepare the reference substance solution and the test solution. The weighed amounts of the test substance are 0.4 g (80%), 0.5 g (100%), and 0.6 g (120%) respectively. Weigh accurately and prepare three concentrations of the test solution. Prepare 2 replicates for each concentration, and a total of 6 test solutions are prepared. Accurately pipette 10 μl of the reference substance solution and 10 μl of the test solution respectively, inject them into the liquid chromatograph, and determine. The results are shown in Table 10.

[0125] Table 10 Range

[0126]

[0127] The results showed that the RSD of danshensu content in 6 test solutions was 0.4%, the RSD of protocatechuic aldehyde content was 2.0%, and the RSD of salvianolic acid B content was 1.3%. The method had good range.

[0128] 2.2.6 Durability

[0129] According to the test method in 2.1, prepare the test solution, place it at room temperature, and accurately pipette 10 μl at 0, 2, 4, 6, 8, 10, 12, and 14 h respectively, inject it into the liquid chromatograph, and determine. The results are shown in Table 11.

[0130] Table 11 Peak Area

[0131]

[0132]

[0133] The results showed that the RSD of the peak area of sodium danshensu measured at each time point was 0.2%, the RSD of the peak area of protocatechuic aldehyde was 0.7%, and the RSD of the peak area of salvianolic acid B was 0.2%. The durability of this method was good.

[0134] Example 3

[0135] Three batches (250201, 250202, 250203) of Guanxinjing tablets were selected, with 2 samples for each batch. According to the detection methods in 1.1 and 2.1, it was detected whether paeoniflorin was contained in Guanxinjing tablets, and the contents of sodium danshensu, protocatechuic aldehyde and salvianolic acid B in Guanxinjing tablets were determined.

[0136] As a result, at the corresponding positions of the chromatograms of the 3 batches of Guanxinjing tablet samples, the reference crude drug chromatogram and the reference substance chromatogram, spots of the same color appeared.

[0137] The contents of sodium danshensu in the 3 batches of Guanxinjing tablets were 0.7726 - 0.8212 mg / tablet, the contents of protocatechuic aldehyde were 0.0749 - 0.1064 mg / tablet, and the contents of salvianolic acid B were 1.9105 - 2.5379 mg / tablet.

[0138] As can be seen from the above examples, the present invention provides a detection method for Guanxinjing tablets. The detection method of the present invention includes (1) identifying paeoniflorin in Guanxinjing tablets by thin layer chromatography, and (2) detecting the contents of sodium danshensu, protocatechuic aldehyde and salvianolic acid B in Guanxinjing tablets by high performance liquid chromatography, filling the methods in the prior art that do not disclose how to detect the contents of each drug component in the monarch drug danshen in Guanxinjing tablets and the methods of drug components in the ministerial drugs, and providing a basis for more accurately detecting Guanxinjing tablets. The method for detecting paeoniflorin in Paeonia lactiflora Pall. by thin layer chromatography of the present invention has good specificity and good durability. The high performance liquid chromatography method for detecting the contents of sodium danshensu, protocatechuic aldehyde and salvianolic acid B in danshen has good specificity, the recovery rate is between 95 - 105%, the accuracy is good, the precision is good, the linear relationship is good, the range is good, and the durability is good. It provides a basis for strictly controlling the quality of Guanxinjing tablets.

[0139] The above are only the preferred embodiments of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and modifications can be made, and these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims

1. A detection method for Guanxinjing tablets, where the Guanxinjing tablets include the monarch drug Salvia miltiorrhiza and the minister drug Paeonia lactiflora. It is characterized in that the detection method includes the following steps: (1) Identifying paeoniflorin in Guanxinjing tablets by thin-layer chromatography 1.1 The active ingredients in Guanxinjing tablets and Paeonia lactiflora Pall. were extracted separately using the same method, prepared into solutions, and the test solution and the control crude drug solution were obtained. 1.2 The paeoniflorin reference substance was mixed with methanol to obtain the reference substance solution. 1.3 The test solution, the control crude drug solution, and the reference substance solution were spotted on the same silica gel G thin-layer plate respectively, developed using the developing agent, taken out, dried, sprayed with the color-developing agent, and heated for color development; whether the test solution contained paeoniflorin was judged according to the color-developing position and color of the control crude drug solution and the reference substance solution. The paeoniflorin in Guanxinjing tablets belongs to Paeonia lactiflora Pall. (2) Determination of the contents of danshensu, protocatechuic aldehyde, and salvianolic acid B in Guanxinjing tablets by high performance liquid chromatography 2.1 The reference substances of danshensu sodium, protocatechuic aldehyde, and salvianolic acid B were mixed with 65 - 75 vt% methanol to obtain the reference substance solution. 2.2 The active ingredients in Guanxinjing tablets were extracted and prepared into a solution to obtain the test solution. 2.3 The reference substance solution and the test solution were respectively injected into the liquid chromatograph for determination. The chromatographic conditions were as follows: the chromatographic column was a C18 column (4.6 mm × 250 mm, 5 μm); acetonitrile was used as mobile phase A, and 0.05 - 0.15% trifluoroacetic acid was used as mobile phase B, the column temperature was 28 - 32 °C, the detection wavelength was 260 - 290 nm, and the number of theoretical plates calculated by the salvianolic acid B peak was not less than 30000. The elution conditions are as follows: from 0 to 20 min, 5% mobile phase A and 95% mobile phase B; from 20 to 30 min, 5 → 9% mobile phase A and 95 → 91% mobile phase B; from 30 to 35 min, 9% mobile phase A and 91% mobile phase B; from 35.1 to 95 min, 20% mobile phase A and 80% mobile phase B; Danshensu, protocatechuic aldehyde, and salvianolic acid B in Guanxinjing tablets belong to Salvia miltiorrhiza Bge.

2. The detection method according to claim 1, wherein The method for extracting the active ingredients from Guanxinjing tablets and Paeonia lactiflora Pall. in step 1.1 was as follows: Guanxinjing tablets or Paeonia lactiflora Pall. were crushed to obtain a powder; the powder was mixed with methanol and sonicated for 25 - 35 min, filtered, and the filtrate was taken; the filtrate was concentrated to obtain a concentrated solution; the concentrated solution was mixed with neutral alumina, evaporated to dryness to obtain a dried product; the dried product was added to a neutral alumina column, eluted, and the eluate was collected to obtain the active ingredient. The active ingredient was evaporated to dryness to obtain a residue, and the residue was dissolved in methanol to respectively prepare the test solution and the control crude drug solution. The mass-to-volume ratio of the powder mixed with methanol was 1 - 1.5 g:25 mL. The mass-to-volume ratio of neutral alumina mixed with the concentrated solution was 0.8 - 1.2 g:0.8 - 1.2 mL. The particle size of the neutral alumina was 200 - 300 mesh. The neutral alumina column contained neutral alumina with a particle size of 200 - 300 mesh, and the inner diameter of the neutral alumina column was 0.8 - 1.2 cm. The solution used for elution was methanol.

3. The detection method according to claim 2, wherein, The mass-to-volume ratio of the paeoniflorin reference substance mixed with methanol in step 1.2 was 0.8 - 1.2 mg:0.8 - 1.2 mL.

4. The detection method according to claim 3, wherein The developing agent described in step 1.3 included components with the following volume ratio: Chloroform:methanol:water was 38 - 42:8 - 12:

1. The color-developing agent described in step 1.3 was a sulfuric acid - ethanol solution; the concentration of sulfuric acid contained in the sulfuric acid - ethanol solution was 10 vt%. The temperature for heating color development in step 1.3 was 100 - 110 °C.

5. The detection method according to claim 4, wherein The concentration of danshensu sodium in the reference substance solution in Step 2.1 is 0.1 - 0.14 mg / mL, the concentration of protocatechuic aldehyde is 10 - 14 μg / mL, and the concentration of salvianolic acid B is 0.26 - 0.30 mg / mL.

6. The detection method according to claim 5, wherein The method for extracting the active ingredients in Guanxinjing tablets in Step 2.2 is as follows: Crush the Guanxinjing tablets to obtain powder; mix the powder with methanol to obtain a mixture, and weigh the mixture; heat the mixture under reflux for 25 - 35 min, cool to obtain the active ingredients; Weigh the obtained active ingredients; make up the weight of the active ingredients to the same as the weight of the mixture with methanol, filter, and take the filtrate to obtain the test solution; The mass - volume ratio of the powder to methanol for mixing is 0.4 - 0.6 g:25 mL.

7. Use of the detection method according to any one of claims 1 - 6 in accurately identifying whether paeoniflorin is contained in Guanxinjing tablets.

8. Use of the detection method according to any one of claims 1 - 6 in accurately detecting the danshensu content in Guanxinjing tablets.

9. Use of the detection method according to any one of claims 1 - 6 in accurately detecting the protocatechuic aldehyde content in Guanxinjing tablets.

10. Use of the detection method according to any one of claims 1 - 6 in accurately detecting the salvianolic acid B content in Guanxinjing tablets.

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